Correlation between antibiotic consumption and the occurrence of multidrug-resistant organisms in a Malaysian tertiary hospital: a 3-year observational study

Inappropriate use of antibiotics has been shown to contribute to the occurrence of multidrug-resistant organisms (MROs). A surveillance study was performed in the largest tertiary care hospital in Kuala Lumpur, Malaysia, from 2018 to 2020 to observe the trends of broad-spectrum antibiotics (beta-lactam/beta-lactamases inhibitors (BL/BLI), extended-spectrum cephalosporins (ESC), and fluoroquinolones (FQ)) and antibiotics against MRO (carbapenems, polymyxins, and glycopeptides) usage and the correlation between antibiotic consumption and MROs. The correlation between 3-year trends of antibiotic consumption (defined daily dose (DDD)/100 admissions) with MRO infection cases (per 100 admissions) was determined using a Jonckheere-Terpstra test and a Pearson’s Correlation coefficient. The antimicrobial resistance trend demonstrated a positive correlation between ESC and FQ towards the development of methicillin-resistant Staphylococcus aureus (MRSA), extended-spectrum beta-lactamase (ESBL)-producing Klebsiella spp, ESBL-producing Escherichia coli (E. coli), and MRO Acinetobacter baumannii (A. baumannii). Increasing carbapenem consumption was positively correlated with the occurrence of ESBL-producing Klebsiella spp and E. coli. Polymyxin use was positively correlated with ESBL-producing Klebsiella spp, MRO A. baumannii, and carbapenem-resistant Enterobacteriaceae. The findings reinforced concerns regarding the association between MRO development, especially with a surge in ESC and FQ consumption. Stricter use of antimicrobials is thus crucial to minimise the risk of emerging resistant organisms.

www.nature.com/scientificreports/ surveillance of antibiotic use and resistance at the local and national levels is vital to evaluate the effectiveness of ASPs and devise new strategies to address the gaps in the existing practices 11,12 . Although the Malaysian national data on antibiotic use and susceptibility/resistance patterns can be found in the annual National Surveillance of Antibiotic Resistance reports, Malaysian Statistics of Medicine, and National Antibiotic Guidelines (NAG), the antibiotic consumption and susceptibility profiles may vary in different localities 13,14 . There is also a lack of evidence available in Malaysia that highlights the influence of broad-spectrum antibiotic usage toward the occurrence of MROs.
To address these existing gaps, the present study evaluated the 3-year trend of broad-spectrum antibiotics and antibiotics against MRO usage in Hospital Kuala Lumpur (HKL) and its influence on the occurrence of MROs. The main objectives of this study are to: (1) evaluate the trend of use of broad-spectrum antibiotics, (2) analyse the trend of occurrence of MROs and (3) determine the influence of broad-spectrum antibiotic usage on the occurrence of MROs. It was hypothesised that an increase in the trend of broad-spectrum antibiotics usage correlates with a surge in the occurrence of MROs.

Materials and methods
Study design and period. Quantitative research was conducted to analyse the three-year trend of systemic broad-spectrum antibiotics use and its influence on the MRO infection rates in HKL, a tertiary care hospital in Kuala Lumpur, Malaysia. It has 73 wards and 1570 beds with 19 clinical departments. Data on antibiotic consumption (expressed through DDD/100 admissions) was obtained from the pharmacy department, while MRO data were retrieved from the infection control department. The protocol was approved by the Medical Research Ethics Committee (MREC), Ministry of Health, Malaysia (NMRR-20-787-54141) and the Clinical Research Committee of HKL and in compliance with ethical principles outlined in the Declaration of Helsinki and Malaysian Good Clinical Practice (GCP) Guideline. The requirement for written informed consent from patients was waived by MREC due to the retrospective nature of the study. Furthermore, precautions were taken to ensure that all data and records confidentiality were only used for this study.
Study definition. Antibiotic. Six groups of antibiotics that are classified under J01 (antibacterial for systemic use) according to the Anatomical Therapeutic Chemical (ATC) Classification System were included in this study. Broad-spectrum antibiotics selected for this study include amoxicillin/clavulanate, ampicillin/sulbactam, piperacillin/tazobactam, ceftriaxone, cefotaxime, ceftazidime, cefoperazone, cefoperazone/sulbactam, cefepime, ciprofloxacin and levofloxacin. Meanwhile, antibiotics against multidrug-resistant organisms (MROs) comprised of imipenem, meropenem, ertapenem, colistin, polymyxin B and vancomycin. Extended-spectrum cephalosporin (ESC) are cephalosporins with better activity against gram-negative microbes than first-generation agents. The following formula was used to calculate the defined daily dose (DDD) per 100 admissions (expressed as DDD/100 admissions) 15 : Rate of MRO infections. The types of MROs analysed in this study were Methicillin-resistant Staphylococcus aureus (MRSA), extended-spectrum beta-lactamase (ESBL)-producing Klebsiella spp, ESBL-producing Escherichia coli, multidrug-resistant organisms (MRO) Acinetobacter baumannii, carbapenem-resistant Enterobacteriaceae (CRE) and vancomycin-resistant enterococci (VRE). The MRO cases were followed up by Infection Control Team through discussions with primary prescribers to differentiate between infection and coloniser; colonisers data were then excluded from this study. The rate of MRO infection per 100 admissions was calculated using the following formula: Statistical analyses. Firstly, the Jonckheere-Terpstra test was performed to determine the trend of monthly antibiotic consumption, the proportion of pathogens and antimicrobial resistance rate over time. The J-T test is a non-parametric method to evaluate the presence of definite trend/order in the data. Separately, to evaluate the presence of linear relationship, Pearson's correlation coefficient was used to investigate the association between antibiotic consumption and bacterial resistance rates. Statistical significance was defined at p < 0.05. Data analyses were conducted using Statistical Package for the Social Sciences (SPSS) version 28.0 (IBM Corporation, Armonk, NY).

Patient involvement.
No patients were involved in this study. There was no plan to disseminate the results to the relevant patient community.

Number of DDD per year =
Total antibiotic usage g for in − patient adults in a year DDD (from WHO)
The top five most frequently prescribed antibiotics, which accounted for 77.8% of all antibiotic usage, were amoxicillin-clavulanate, ampicillin-sulbactam, piperacillin-tazobactam, ceftriaxone and cefepime. All the antibiotics exhibited increasing use throughout the study period except for ampicillin-sulbactam and piperacillintazobactam, which demonstrated a dip in 2019 and 2020. Consumption of amoxicillin-clavulanate, ceftriaxone, cefoperazone, cefepime, meropenem and ertapenem was significantly greater (p < 0.05) throughout the 3-year observation.

Correlation between antibiotic consumption and MRO infection rates. The correlation between
antibiotic consumption and MRO occurrence is summarised in Table 3

Discussion
The study reported the antibiotic usage and antimicrobial resistance patterns in the biggest tertiary hospital in Kuala Lumpur, Malaysia. Antibiotic Point Prevalent Survey of HKL in 2018 revealed that 88.5% of antibiotics prescribed was for empirical therapies 16 . Jamaluddin et al. reported that empirical antimicrobials accounted for approximately 65% of prescribed antibiotics, notably throughout medical, surgical, orthopaedic and oncology wards in Malaysia 17 .The initial antimicrobial therapy is based on clinical judgments, emphasising likely pathogens while awaiting laboratory reports. Ideally, causative microorganisms' identification enables the clinicians to select the most appropriate antibiotic, which improves overall patient outcomes 18 . Nevertheless, the culture and sensitivity results generally take more than 48 h to be available. Hence, the clinicians had to rely on the most likely organisms and site of infections for antibiotic selection 17 .
In this study, the most consumed antibiotics for hospitalised patients were broad-spectrum antibiotics subgroups, ESC and BL/BLI, comprising 86.4% of total antibiotic consumption. These results are comparable across Ministry of Health (MOH) hospitals, university hospitals, military hospitals, and private hospitals in Malaysia 14 . Similarly, the reports from the Global Point prevalence study (PPS) revealed BLI and ESC are among the top three antibiotics prescribed worldwide 18 . Furthermore, the total utilisation of BL/BLI and ESC increased steadily every year, where a substantial increase was evident from 2019 to 2020 by 26.5% and 39.7%, respectively. Meanwhile, a significant rise in carbapenem, mainly ertapenem, was observed in this study. This might be related to the efforts of de-escalating broad-spectrum carbapenem such as imipenem or meropenem to ertapenem in ESBL infection. However, meropenem consumption increased steadily over the years as a preferred choice of empirical carbapenem due to its safety profile and convenient dosing.
Implementation of Protocol for Antimicrobial Stewardship (AMS) by MOH in 2014 was a successful approach to reduce unnecessary and inappropriate use of antibiotics, resulting in the decreasing trend of total antibiotic consumption in MOH hospitals 13 . However, there was a hike in HKL's entire antibiotic consumption trend, especially in 2020. Given that, Haug et al. suggested a few factors that can influence the variability in antibiotic consumption, including higher nurse staffing and larger proportions of shorter (< 2 days) and longer (> 10 days) hospital stays can lead to an increased antibiotic use 19 . Moreover, variations in local antimicrobial resistance patterns, physicians' prescribing behaviour and workloads can also lead to discrepancies in antibiotic consumption 20,21 . Therefore, it is essential to consider these factors when comparing antibiotic utilisation with the national reports.  23 . In the present study, MRSA and ESBLs showed increased resistance patterns throughout the 3-year observation period. The increase in infection rate may result from high usage of certain antibiotic groups such as FQ and ESC and possibly poor infection control. A major part of healthcare-associated infections is avoidable through effective infection prevention and control strategy that will substantially contain antimicrobial resistance and outbreaks of highly transmissible diseases, including the COVID-19 pandemic. It is evident that infection rate can be contained by establishing local and national standard operating procedures (SOP), especially on five major components that include: (i) modification of system to facilitate good practice, (ii) training and educating healthcare personnel, (iii) observing health practices, procedures and outcomes and delivering timely feedback, (iv) refining healthcare message, and (v) adapting to a safety climate 24 . A study by Nori et al. on SARS-CoV-2 identified the most common opportunistic pathogens were S. aureus (44%), P. aeruginosa (16%), Klebsiella spp. (10%), Enterobacter. spp. (8%) and E. coli (4%) while MRO isolates were present in 15% of the testing 25 . The emergence of MRO CRE poses challenges to healthcare institutions globally, including Malaysia, since carbapenem is one of the 'last-line antimicrobials Table 3. Correlation between antibiotic consumption and MRO occurrence. BL/BLIs = beta lactam/betalactamase inhibitors, ESC = extended-spectrum cephalosporin (third generation and fourth generation cephalosporins), FQ = fluoroquinolones. Significant result *p < 0.05, **p < 0.01.

Organism
Antibiotic group www.nature.com/scientificreports/ where treatment options are limited, and higher mortality rates are reported compared to other MRO strains 26 . Moreover, the review of carbapenem at 72 h following AMS is believed to assist in carbapenems de-escalation to substantially reduce the risk of CRE 27 . Antibiotic resistance directly influences antimicrobial prescription by increasing the burden by mitigating certain classes of antibiotic class to another to reduce the selective pressure towards a particular antimicrobial class. Based on the current investigation, ESC and FQ had a stronger association with MRO than other antibiotic groups, including MRSA, ESBLs and MRO A. baumannii. Several studies described that ESC-resistant isolates were associated with gram-negative microorganisms' production of ESBL, plasmid-mediated AmpCs, carbapenemase enzymes and MRSA occurrence 28,29 . In addition, Wushouer et al. reported significant positive correlations between MRSA infection rates and the consumption of third generation cephalosporins, carbapenems and glycopeptides 22 .
FQ consumption contributed to the prevalence of ESBL-producing pathogens, consistent with a surveillance study in China 30 . Ryu et al. explained that the positive correlation between FQ and Klebsiella spp was related to the transferable plasmid (quinolone-resistant (qnr) gene) in ESBL-producing strains that lead to FQ resistance 31 . Klebsiella spp, especially K. pneumoniae, adapts well in the hospital environment and survives longer than other Enterobacteriaceae; thus, allowing for easier cross-infection, which deems for stricter control of the antibiotics in the future 32 . In the current study, carbapenem use showed a positive correlation with ESBL infections despite being used in the AMS strategy as the definitive treatment for ESBLs. Martìnez et al. reported that previous carbapenem use was a significant risk factor for ESBL-Klebsiella spp. and E. coli based on the possibilities of antibiotic enriching the gut reservoir with ESBL in patients already being colonised with a low number of microbes or promoting bacterial settlement following transmission event 33 . Besides, the positive correlation between polymyxins and ESBL-Klebsiella spp, MRO A. baumannii, and CRE may be linked to the chromosomal mutations of the carbapenem-resistant A. baumannii ST1 and CrrB single nucleotide polymorphism (SNP) mutants of polymyxins 34,35 .
The negative incidence of VRE to glycopeptides in the study may be linked to the comprehensive strategies that include appropriate dosing and duration of administration 24 . Interestingly, a negative correlation between BL/BLI and MRO A. Baumannii and CRE may have been contributed by reversed bacterial resistance mechanisms based on the antibiotic's preferential targeting of specific cell wall assembly machinery 36 .
In order to curb the increasing antibiotic consumption trend, we have introduced several actions plans to improve antibiotic usage in our setting, including strict implementation of antibiotic request forms and antibiotic stop orders. The prescribers are required to submit an antibiotic request form upfront when prescribing six types of antibiotics, including meropenem, imipenem, ertapenem, vancomycin, colistin and polymyxin B. An antibiotic stop order is activated if these antibiotics are continued beyond 72 h, without any justification or evidence from culture and sensitivity results. In addition, our setting also embarked on persuasive antimicrobial stewardship initiatives such as education to all healthcare professionals through antimicrobial awareness week events and antimicrobial stewardship grand ward rounds.
Limitations. The study focused solely on the prevalence of antibiotic usage, primarily due to the nature of the cross-sectional study design. Other factors that could potentially influence antimicrobial use include the COVID-19 outbreak in 2020 and the patient case-mix. Therefore, the retrospective and observational study design cannot confirm the causal relationship between antibiotics consumption and MROs infection rates. Nevertheless, there was a positive correlation between the two variables. Therefore, the findings can suggest ways to reduce antimicrobial resistance. Furthermore, due to the different dosage requirements, antibiotic consumption was measured using DDD, which may be inaccurate and underestimates certain patient groups such as renal impairment. Therefore, it may not represent the prescribed daily dose for all patients.
Moreover, there was a lack of patient-specific data for detailed risk factor evaluation. Since certain patients with a higher risk of MRO infection at baseline or cross-infection of MROs may affect the statistical significance, future research should include demographic data to address this gap. In addition, more data points (eg.10-year trends) should be considered to prove the correlation between antibiotics' consumption and MROs infection.

Conclusion
The overall data presented a fair indicator for antibiotic use and resistance pattern in a tertiary hospital in Malaysia. Broad-spectrum antibiotics and antibiotics against MRO consumption increased significantly throughout the study, while the antimicrobial resistance trend in the hospital decreased, except for MRSA and ESBL. In addition, ESC and FQ consumption was positively correlated with the emergence of MRSA, ESBL-producing Klebsiella spp, ESBL-producing E. coli and MRO A. baumannii resistance. Continuous review and reinforcement of AMS in tertiary care settings are thus crucial to overcome the potential threats by MROs.